β-1,3-glucan synthesis

Ibrexafungerp is being developed as the first oral and IV GSI (Intravenous glucan synthase inhibitor) for the treatment and prevention of fungal infections, including serious and life-threatening infections due to Candida spp., Aspergillus spp., and Pneumocystis jirovecii, with the potential to provide the therapeutic advantages of both IV and oral formulations. Ibrexafungerp causes a decrease in (1→3)-β-D-glucan polymers and a weakening of the fungal cell wall. Ibrexafungerp is structurally distinct from echinocandins and interacts differently with the target enzyme (Figure 2). Although the binding site on (1→3)-β-D-glucan synthase for ibrexafungerp partially overlaps with a binding site for echinocandins, it appears to be nonidentical, resulting in a lower rate of resistance to ibrexafungerp. In in vitro studies, ibrexafungerp activity against wild-type and echinocandin-resistant strains of Candida spp. in the presence of fks mutations was minimally affected. Thus, ibrexafungerp has limited potential for cross-resistance with echinocandins. [1]
Ibrexafungerp demonstrates broad in vitro activity against a range of Aspergillus spp. isolates and Candida isolates, including C. glabrata and C. auris, which exhibit fks1 and fks2 point mutations associated with resistance to echinocandin antifungals. Among Candida species with reduced fluconazole susceptibility, including C. glabrata, C. krusei, C. tropicalis, and C. parapsilosis, MIC50 ranges with ibrexafungerp were 0.125–1 μg/mL, 0.5–1 μg/mL, <0.03–1 μg/mL, and 0.25–1 μg/mL, respectively. Additionally, as reported by Zhu [32] using isolates obtained from New York patients, the in vitro activity against C. auris of ibrexafungerp (ranging from 0.05 to 0.5 μg/mL) was superior to that of fluconazole (ranging from 2 to >256 μg/mL), and comparable or superior to that of echinocandins (ranging from 0.015 to 16 μg/mL). This observation was confirmed by other studies using global strains. Ibrexafungerp showed a wild-type MIC distribution against ~80% of echinocandin-resistant Candida spp. isolates tested, suggesting that fks mutations have less of an effect on the in vitro activity of ibrexafungerp. [1]

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Ibrexafungerp (SCY-078) is a novel first-in-class antifungal agent targeting glucan synthase. Candida auris is an emerging multidrug-resistant species that has caused outbreaks on five continents. We investigated the in vitro activity of ibrexafungerp against C. auris by applying EUCAST E.Def 7.3.1 methodology. C. albicans and C. glabrata, as well as anidulafungin, micafungin, amphotericin B, fluconazole, voriconazole, and isavuconazole, were included as comparators. Three C. auris reference strains (CBS12372, CBS12373, and CBS10913) and 122 C. auris, 16 C. albicans, and 16 C. glabrata isolates were evaluated. C. albicans ATCC 64548, C. parapsilosis ATCC 22019, and C. krusei ATCC 6258 served as quality control strains. Echinocandin-resistant isolates were fks sequenced. MIC ranges and modal MIC and MIC50 values were determined. Wild-type upper limits (the upper MIC value where the wild-type distribution ends) were determined according to EUCAST principles for setting ECOFFs. Nine repetitions of three QC strains and MICs for C. albicans and C. glabrata yielded narrow MIC ranges with modal MICs in agreement with established EUCAST modal MICs, confirming a robust test performance. The ibrexafungerp MICs against C. auris isolates displayed a Gaussian distribution with a modal MIC (range) of 0.5 mg/liter (0.06 to 2 mg/liter), suggesting uniform susceptibility. Of 122 isolates, 8 were echinocandin resistant and harbored the S639F Fks1 alteration. All but one were fluconazole resistant, and the MIC distributions for voriconazole and isavuconazole were multimodal confirming variable susceptibility. Ibrexafungerp demonstrated promising activity against C. auris, including isolates resistant to echinocandins and/or other agents. The MICs were similar to those reported for the Clinical and Laboratory Standards Institute method, suggesting that a common clinical breakpoint may be appropriate [4].

SCY-078 (MK-3118) is a novel, semisynthetic derivative of enfumafungin and represents the first compound of the triterpene class of antifungals. SCY-078 exhibits potent inhibition of β-(1,3)-d-glucan synthesis, an essential cell wall component of many pathogenic fungi, including Candida spp. and Aspergillus spp. SCY-078 is currently in phase 2 clinical development for the treatment of invasive fungal diseases. In vitro disposition studies to assess solubility, intestinal permeability, and metabolic stability were predictive of good oral bioavailability. Preclinical pharmacokinetic studies were consistent with once-daily administration to humans. After intravenous delivery, plasma clearance in rodents and dogs was low, representing <15% and <25% of hepatic blood flow, respectively. The terminal elimination-phase half-life was 5.5 to 8.7 h in rodents, and it was ∼9.3 h in dogs. The volume of distribution at steady-state was high (4.7 to 5.3 liters/kg), a finding suggestive of extensive tissue distribution. Exposure of SCY-078 in kidney tissue, a target organ for invasive fungal disease such as candidiasis, exceeded plasma by 20- to 25-fold for the area under the concentration-time curve from 0 h to infinity (AUC0-∞) and Cmax SCY-078 achieved efficacy endpoints following oral delivery across multiple murine models of disseminated candidiasis. The pharmacokinetic/pharmacodynamic indices Cmax/MIC and AUC/MIC correlated with outcome. Target therapeutic exposure, expressed as the plasma AUC0-24, was comparable across models, with an upper value of 11.2 μg·h/ml (15.4 μM·h); the corresponding mean value for free drug AUC/MIC was ∼0.75. Overall, these results demonstrate that SCY-078 has the oral and intravenous (i.v.) pharmacokinetic properties and potency in murine infection models of disseminated candidiasis to support further investigation as a novel i.v. and oral treatment for invasive fungal diseases.[2]

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Candida auris has been shown to have a high risk of skin colonization in hospitalized patients, possibly contributing to nosocomial spread. In a guinea pig skin model, animals were evaluated for clinical appearance, tissue fungal burden, histology, and pharmacokinetics. Oral dosing with 10 mg/kg ibrexafungerp (IBX) reduced the severity of lesions and significantly reduced the C. auris fungal burden in infected animals compared with untreated controls. This indicates promise for use of IBX in controlling skin infection and colonization of hospitalized patients [3].

Solubility.[2]
The solubility of IbrexafungerpSCY-078 was measured in simulated gastric fluid (SGF), fasted-state simulated intestinal fluid (FaSSIF), and fed-state intestinal fluid (FeSSIF) by Crystal Pharmatech. In general, approximately 15 mg of solid was weighed into a 4-ml vial, 3.0 ml of medium was added, and the suspensions were stirred on a rolling incubator (25 rpm) at an ambient room temperature for 24 h. After incubation, 0.5 ml of suspension was centrifuged and filtered (0.45-μm pore size), and the concentration of SCY-078 was determined in the supernatant by high-pressure liquid chromatography (HPLC) with UV detection. The quantity of SCY-078 and the volume of medium was adjusted as required to determine the solubility in each medium.

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In vitro metabolic stability in hepatic microsomes.[2]
The metabolic stability of IbrexafungerpSCY-078 was evaluated with male and female mouse, male rat, male and female dog, and mixed-gender human liver microsomes. SCY-078 (1 μM) was incubated with pooled liver microsomes (0.5 mg protein/ml) for 0, 5, 10, 20, and 30 min at 37°C in the presence of NADPH. Aliquots were taken at each sampling time point and extracted with 5 volumes of ice-cold acetonitrile containing the internal standard (d9-SCY-078, 125 ng/ml). Supernatants from the incubation mixtures were analyzed for the parent compound by LC-MS/MS according to the conditions described below. The metabolic competency of microsomal preparations was established using the control compounds 7-ethoxycoumarin, propranolol, and verapamil. The in vitro intrinsic clearance (CLint), the CLint scaled to the in vivo intrinsic clearance (CL′int), and the half-life were determined for SCY-078 incubated with microsomes from each species.

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In vitro plasma protein binding and blood distribution.[2]
Protein binding of [3H]SCY-078 in mouse, rat, dog, and human plasma was determined by means of equilibrium dialysis and liquid scintillation counting. Tritium-labeled SCY-078 in ethanol was added to unlabeled SCY-078 in methanol to prepare stock solutions at 10, 100, and 1,000 μM Ibrexafungerp/SCY-078 with constant activity. Protein binding was determined against isotonic phosphate-buffered saline (pH 7.4) at 37°C for 24 h. Aliquots (10 μl) of each stock solution were added to pooled DBA mouse (n = >10 animals), Sprague-Dawley rat, beagle dog, or human plasma (1 ml) to achieve final total concentrations of 0.1, 1, and 10 μM Ibrexafungerp/SCY-078. The final concentration of the organic solvent was 0.9% (vol/vol). Dialysis was performed in 1-ml acrylic dialysis cells, where plasma and buffer were separated by a 12.4-kDa cutoff membrane (Sigma, St. Louis, MO). After incubation, aliquots (100 μl) of plasma and phosphate buffer were counted for radioactivity, and unbound fractions of SCY-078 were estimated based on the following calculation: unbound fraction = radioactivity (dpm/0.1 ml) in buffer/total radioactivity (dpm/0.1 ml) in plasma.

In vitro permeability through Caco2 cell monolayers.[2]
Caco2 cells (ATCC CRL-2102) were cultured in Dulbecco modified Eagle medium with the dipeptide form of l-glutamine (GlutaMAX), 10% (vol/vol) fetal bovine serum, and 1% (vol/vol) penicillin-streptomycin at 10,000 U/ml in a 75-ml flask at 37°C in a humidified atmosphere of 5% CO2. Near-confluent Caco-2 cell cultures were harvested by trypsinization with 0.25% trypsin at 37°C for 5 min and resuspended in culture medium. The cells were seeded onto semipermeable filter inserts at a density of approximately 200,000 cells/cm2. The cell culture medium was changed every 2 to 3 days over a total of 21 days of culture. On the day of the assay, the cell monolayers were rinsed with transport medium (Hanks balanced salt solution with 25 mM glucose and 25 mM HEPES), and the absorptive permeation of Ibrexafungerp/SCY-078 was evaluated by measuring the flux from the apical to the basolateral compartments. Cell monolayers were incubated with Ibrexafungerp/SCY-078 (5 μM) in triplicate for 2 h at 37°C. Samples were removed from the apical and basolateral compartments after incubation and assayed for test compound concentrations by LC-MS/MS. The apparent permeability coefficient (Papp; cm/s) was calculated as follows: Papp = 1/A·C0 (dQ/dt), where dQ/dt is the rate of drug appearance in the basolateral compartment (μmol/s), C0 is the initial drug concentration in the donor compartment (μM), and A is the surface area of the monolayer (cm2). The results were expressed as the mean ± the SD from triplicate samples (n = 3).

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Susceptibility testing.[4]
EUCAST MICs were determined following E.Def 7.3.1 methodology. Ibrexafungerp (SCY-078) pure substance was stored in aliquots at −80°C, and stock solutions were prepared in dimethyl sulfoxide (5,000 mg/liter). The final drug concentration ranges studied were 0.008 to 4 mg/liter. The following comparator compounds were also investigated (source of compound with the final concentration ranges in parentheses): anidulafungin (0.004 to 4 mg/liter for C. albicans and C. glabrata isolates and 0.03 to 32 mg/liter for C. auris), micafungin (0.004 to 4 mg/liter for C. albicans and C. glabrata isolates and 0.03 to 32 mg/liter for C. auris), amphotericin B (0.004 to 4 mg/liter), fluconazole (0.03 to 32 mg/liter for bloodstream isolates and 0.5 to 256 mg/liter for C. auris), isavuconazole (0.004 to 4 mg/liter), and voriconazole (0.004 to 4 mg/liter). Cell culture-treated samples were used throughout. Microtiter plates with 2-fold dilutions were prepared and frozen at −80°C prior to use.

Murine models of disseminated candidiasis. [2]
The in vivo activity of SCY-078 was evaluated using two murine models of disseminated candidiasis to establish the pharmacokinetic exposure target and PK/PD measures associated with efficacy. In the first model, the target therapeutic exposure was established across four independent experiments, each employing 7 days of twice-daily (BID) oral treatment initiated shortly after infection. To establish the PK/PD parameters associated with the outcome, the efficacy of SCY-078 was compared after single or fractionated doses starting 16 h after the infectious challenge. [2]
A disseminated Candida infection was induced in C′5-deficient DBA/2N mice, weighing on average 20 g, by i.v. inoculation with C. albicans MY1055. C. albicans MY1055 was cultured on Sabouraud dextrose agar plates at 35°C for 24 h. Yeast cells were washed from the surface of agar plates into sterile saline, and the cell concentrations were quantitated by using a hemocytometer. Viable cell counts were confirmed by a serial 10-fold dilution of the cell suspension and plating on SDA plates. Plates were incubated for 24 to 48 h at 35°C, whereupon the numbers of CFU were determined. The in vitro activity of SCY-078 against the MY1055 isolate was evaluated using broth microdilution assays as described in CLSI M27-A3. The MIC endpoints were based on 50% inhibition of fungal growth at 24 h. [2]
For infection, 0.2 ml of a blastospore suspension containing between 2.44 × 104 and 3.56 × 104 CFU of C. albicans MY1055 was inoculated into the lateral tail vein. Mice were housed in groups of up to 10 animals in sterile microisolator cages with sterile bedding. Water and food were provided ad lib. The infected and nonmedicated (sham-treated control) animals (n = 20) received vehicle only. Treatment groups comprised five animals each, with an additional three animals included for PK analysis for SCY-078. Blood samples were collected from infected satellite PK mice at typically 0, 0.25, 0.5, 1, 2, 4, 6, and 24 h after dose 13 on day 7. Kidneys from five mice were aseptically removed from each treatment group at day 7 after infectious challenge, unless otherwise indicated. [2]
Therapy with SCY-078, caspofungin or fluconazole was initiated within 15 to 30 min after challenge. Mice were treated with SCY-078 with BID p.o. doses of 6.25, 12.5, or 25 mg/kg administered in a “fit-for-purpose” formulation. Caspofungin was administered twice daily via the intraperitoneal route at doses of 0.0078, 0.03, 0.125, and 0.5 mg/kg. Fluconazole was administered p.o. BID at doses of 0.078, 0.31, 1.25, and 5.0 mg/kg. At day 7 after challenge, the mice (n = 5/group) were euthanized, and both kidneys were aseptically removed, placed in sterile Whirl-Pak bags, weighed, and homogenized in 5 ml of sterile physiological saline. Kidney homogenates were serially 10-fold diluted in sterile saline and plated on SDA. Plates were incubated at 35°C and counted after 30 to 48 h of incubation. The CFU/g of kidney were determined, and counts from treatment groups were compared to counts from sham-treated controls using a paired two-tailed t test (Microsoft Excel). The percent clearance was determined as the number of mice with no detectable yeast, with a limit of detection of 50 yeast cells per pair of kidneys because of the dilution scheme. For data from individual mice where no detectable yeast were recovered from paired kidneys, 9.8 was entered into a Microsoft Excel spreadsheet formula [log10 (5 × raw count)/paired kidney weight)] so that the counts would be one less than the limit of detection or 49 cells per pair of kidneys. [2]
Pharmacokinetic analysis was performed on samples collected from satellite-infected mice (n = 3/group) after SCY-078 dose 13 on treatment day 7. Tail bleeds were obtained (20 μl collected into 60 μl of 0.1 M sodium citrate) at time points from 15 min to 6 h, concluding with a terminal bleed at 24 h. Samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) after protein precipitation. Plasma exposure for SCY-078 was calculated from the in vitro plasma/whole-blood distribution ratio (see below). [2]
Efficacy was determined in a delayed treatment model based on the protocol described above using typically 5 mice per treatment group. For the single delayed-dose treatment model, a single dose of SCY-078 was administered 16 h after infection. The target endpoint for efficacy in this model was a static effect on the tissue burden measured at 96 h posttreatment. Caspofungin and fluconazole were not evaluated in this model. In the fractionated delayed-dose treatment model, SCY-078 was administered as divided doses of either two half doses or four quarter doses relative to the single dose. The total doses were 12.5, 25, and 50 mg/kg administered as a suspension in a “fit-for-purpose” formulation. Divided doses were administered at either 0 or 48 h or at 0, 24, 48, and 72 h relative to the single dose for the half and quarter doses, respectively. Caspofungin and fluconazole were not evaluated in this model.

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SCY-078 pharmacokinetics in plasma and kidney tissue. [2]
The pharmacokinetics and bioavailability of SCY-078 were evaluated in uninfected mouse, rat, and dog after p.o. and i.v. administration. Avastis and MPI Research are Association for Assessment and Accreditation of Laboratory Animal Care International (AAALAC)-approved facilities. Aqueous formulations of SCY-078 were prepared for p.o. or i.v. delivery, 0.45% (mass/vol) saline or 0.45% (mass/vol) saline containing 2% (mass/vol) PEG400 for i.v. administration and administered at either a 4-, 5-, or 5-ml/kg dose for mice, rats, or dogs, respectively. The formulations used for the PK studies were prepared from an optimized SCY-078 salt form that enhanced the fraction absorbed relative to the fit-for-purpose SCY-078 formulations used for the murine efficacy studies. Blood samples for PK studies were collected into tubes containing K2EDTA anticoagulant and stored on wet ice until centrifuged and processed for plasma (stored at approximately −80°C). The peak concentration (Cmax), the time to maximum concentration (Tmax), the half-life, and the AUC were determined from composite mean plasma concentration-time data for rodents and individual plasma concentration-time data for dogs. All doses and plasma concentrations of SCY-078 are presented as free base.

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Mice. [2]
Female CD-1 mice (n = 5/sex/time point/group) weighing approximately 25 g were administered SCY-078 by oral gavage either once daily or as multiple twice-per-day (Q12h) treatments, with individual doses ranging between 3 and 100 mg/kg (6 to 150 mg/kg/day). Mice also received a single 1-mg/ml i.v. dose of SCY-078. Plasma specimens were collected prior to dosing and at 0.25, 0.5, 1, 2, 4, 6, 9, 12, 18, 24, 36, 48, and 60 h after the single i.v. dose and at the same time points from 1 h postdose onward following oral dose 13 on day 7 of treatment. On the first day of oral treatment, plasma specimens were collected immediately before each of the two doses and 1, 2, 4, 6, 9, and 12 h postdosing. Two additional samples (predose and at approximately Tmax) were collected after the first dose on day 4 of treatment to determine whether exposure had reached steady-state levels. Kidney tissues were collected at 2, 4, 12, 24, 36, 48, and 60 h after oral dose 13 on treatment day 7, blotted dry, and stored at approximately −80°C until analysis.

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Rats. [2]
Male and female Wistar Han rats (n = 3/sex/group/time point) weighing 0.329 to 0.398 kg (males) or 0.187 to 0.244 kg (females) received a single 5-mg/kg dose of SCY-078 as an i.v. injection in the lateral tail administered over 5 min or an oral dose of 20 mg/kg as described above. Blood specimens were collected from four animals/time point/group using a sparse sampling strategy via the sublingual vein at time intervals to 72 h postdosing and processed as described above.

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Dogs. [2]
Treatment naive male and female beagle dogs (n = 3/sex/group/time point) weighing 8 to 12 kg (males) or 6 to 10 kg (females) were given a single 5-mg/kg dose of SCY-078 as an i.v. injection over 5 min or an oral dose of 20 mg/kg as described above. Blood specimens were collected from the jugular vein at various time intervals to 72 h postdosing and processed as described above.

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Analysis of SCY-078 in biological samples: mouse kidney tissue homogenization. [2]
On the day of analysis, either treatment-naive (drug-free) kidney tissue or freshly thawed tissue samples from PK studies were weighed and homogenized in 3 volumes of phosphate-buffered saline. The density of kidney tissue used for the determination of volume was assumed to be 1.0 g/ml. [2]
Whole-kidney tissue samples were homogenized in parallel by means of a Precellys 24 bead mill homogenizer using 2-ml Precellys tubes containing 1.4-mm-diameter ceramic beads (CK14) and a single 15-s cycle of 5,500 rpm. Parallel bead homogenization increased throughput and avoided potential cross-contamination of samples since each sample was wholly contained and homogenized within its own sealed vial with no direct contact between the homogenizing instrument or other samples.

Absorption, Distribution and Excretion
Ibrexafungerp given at a dose of 300 mg twice daily reaches a Cmax of 435 ng/mL, with a Tmax of 4-6 hours, and an AUC0-24 of 6832 h\*ng/mL.
90% of a radiolabelled oral dose of ibrexafungerp is recovered in the feces, with 51% as the unchanged parent drug. 1% of a radiolabelled oral dose is recovered in the urine.
The volume of distribution at steady state is approximately 600 L.
Clearance values of 53.6 L/h and 56.1 L/h have been reported.

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Metabolism / Metabolites
Ibrexafungerp is hydroxylated by CYP3A4 before glucuronide or sulfate conjugation of the hydroxyl group before elimination.

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Biological Half-Life
The elimination half life of ibrexafungerp is approximately 20 hours.

Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the use of ibrexafungerp during breastfeeding. The drug is over 99% protein bound, so amounts in milk are likely to be very low. If ibrexafungerp is required by the mother of an older infant, it is not a reason to discontinue breastfeeding, but until more data become available, an alternate drug may be preferred, especially while nursing a newborn or preterm infant.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

[1]. Ibrexafungerp: A Novel Oral Triterpenoid Antifungal in Development for the Treatment of Candida auris Infections. Antibiotics (Basel). 2020 Aug 25;9(9):539.
[2]. Preclinical Pharmacokinetics and Pharmacodynamic Target of SCY-078, a First-in-Class Orally Active Antifungal Glucan Synthesis Inhibitor, in Murine Models of Disseminated Candidiasis. Antimicrob Agents Chemother. 2017 Mar 24;61(4):e02068-16.
[3]. Efficacy of Ibrexafungerp (SCY-078) against Candida auris in an In Vivo Guinea Pig Cutaneous Infection Model. Antimicrob Agents Chemother. 2020 Sep 21;64(10):e00854-20.
[4]. In Vitro Activity of Ibrexafungerp (SCY-078) against Candida auris Isolates as Determined by EUCAST Methodology and Comparison with Activity against C. albicans and C. glabrata and with the Activities of Six Comparator Agents. Antimicrob Agents Chemother. 2020 Feb 21;64(3):e02136-19.

See also: Ibrexafungerp (has active moiety).

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Drug Indication
Treatment of invasive candidiasis

C50H75N5O11

922.1574

921.546

C, 65.12; H, 8.20; N, 7.59; O, 19.08

1965291-08-0

1207753-03-4;1965291-08-0 (citrate);

137552087

White to off-white solid powder

6

15

14

66

1650

12

C[C@H](C(C)C)[C@]1(CC[C@@]2([C@H]3CC[C@H]4[C@]5(COC[C@]4(C3=CC[C@]2([C@@H]1C(=O)O)C)C[C@H]([C@@H]5OC[C@@](C)(C(C)(C)C)N)N6C(=NC=N6)C7=CC=NC=C7)C)C)C.C(C(=O)O)C(CC(=O)O)(C(=O)O)O

WKIRTJACGBEXBZ-FQGZCCSZSA-N

InChI=1S/C44H67N5O4.C6H8O7/c1-27(2)28(3)39(7)18-19-41(9)30-12-13-33-40(8)23-52-25-44(33,31(30)14-17-42(41,10)34(39)37(50)51)22-32(35(40)53-24-43(11,45)38(4,5)6)49-36(47-26-48-49)29-15-20-46-21-16-29;7-3(8)1-6(13,5(11)12)2-4(9)10/h14-16,20-21,26-28,30,32-35H,12-13,17-19,22-25,45H2,1-11H3,(H,50,51);13H,1-2H2,(H,7,8)(H,9,10)(H,11,12)/t28-,30+,32-,33+,34-,35+,39-,40-,41-,42+,43+,44+;/m1./s1

(1R,5S,6R,7R,10R,11R,14R,15S,20R,21R)-21-[(2R)-2-amino-2,3,3-trimethylbutoxy]-5,7,10,15-tetramethyl-7-[(2R)-3-methylbutan-2-yl]-20-(5-pyridin-4-yl-1,2,4-triazol-1-yl)-17-oxapentacyclo[13.3.3.01,14.02,11.05,10]henicos-2-ene-6-carboxylic acid;2-hydroxypropane-1,2,3-tricarboxylic acid

Ibrexafungerp citrate; SCY-078 citrate; 1965291-08-0; M4NU2SDX3E; brexafemme;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)